Table Of ContentZootaxa 4642 (1): 001–079 ISSN 1175-5326 (print edition)
Monograph ZOOTAXA
https://www.mapress.com/j/zt/
Copyright © 2019 Magnolia Press
ISSN 1175-5334 (online edition)
https://doi.org/10.11646/zootaxa.4642.1.1
http://zoobank.org/urn:lsid:zoobank.org:pub:42548CEC-E20A-425A-AD8B-5EED9F9DF063
ZOOTAXA
4642
Molecular phylogenetics of snailfishes (Cottoidei: Liparidae) based on
MtDNA and RADseq genomic analyses, with comments on selected
morphological characters
JAMES WILDER ORR1, INGRID SPIES2, DUANE E. STEVENSON3, GARY C. LONGO4,
YOSHIAKI KAI5, SAM GHODS6 & MYLES HOLLOWED7
1NOAA, National Marine Fisheries Service, Alaska Fisheries Science Center, RACE Division, 7600 Sand Point Way NE, Seattle,
Washington, 98115, U.S.A. E-mail: [email protected]
2NOAA, National Marine Fisheries Service, Alaska Fisheries Science Center, REFM Division, 7600 Sand Point Way NE, Seattle,
Washington, 98115, U.S.A. E-mail: [email protected]
3NOAA, National Marine Fisheries Service, Alaska Fisheries Science Center, RACE and FMA Divisions, 7600 Sand Point Way NE,
Seattle, Washington, 98115, U.S.A. E-mail: [email protected]
4NOAA, National Marine Fisheries Service, Northwest Fisheries Science Center, 2725 Montlake Boulevard East, Seattle, Washing-
ton, 98112, U.S.A. E-mail: [email protected]
5Maizuru Fisheries Research Station, Field Science Education and Research Center, Kyoto University, Nagahama, Maizuru, Kyoto
625-0086, Japan. E-mail: [email protected]
6School of Aquatic and Fishery Sciences, and Burke Museum of Natural History and Culture, University of Washington, Seattle,
Washington, 98195, U.S.A. E-mail: [email protected]
7School of Aquatic and Fishery Sciences, and Burke Museum of Natural History and Culture, University of Washington, Seattle,
Washington, 98195, U.S.A. E-mail: [email protected].
Magnolia Press
Auckland, New Zealand
Accepted by E. Hilton: 31 May 2019; published: 22 Jul. 2019
Licensed under a Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0
JAMES WILDER ORR, INGRID SPIES, DUANE E. STEVENSON, GARY C. LONGO, YOSHIAKI KAI,
SAM GHODS & MYLES HOLLOWED
Molecular phylogenetics of snailfishes (Cottoidei: Liparidae) based on MtDNA and RADseq
genomic analyses, with comments on selected morphological characters
(Zootaxa 4642)
79 pp.; 30 cm.
22 Jul. 2019
ISBN 978-1-77670-729-4 (paperback)
ISBN 978-1-77670-730-0 (Online edition)
FIRST PUBLISHED IN 2019 BY
Magnolia Press
P.O. Box 41-383
Auckland 1346
New Zealand
e-mail: [email protected]
https://www.mapress.com/j/zt
© 2019 Magnolia Press
ISSN 1175-5326 (Print edition)
ISSN 1175-5334 (Online edition)
2 · Zootaxa 4642 (1) © 2019 Magnolia Press ORR ET AL.
Table of Contents
Abstract ...................................................................................................3
Introduction ................................................................................................3
Materials and methods ......................................................................................11
Results ..................................................................................................14
Discussion ................................................................................................20
Morphology. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Zoogeography and ecology ...................................................................................40
Conclusions ...............................................................................................42
Acknowledgments .........................................................................................44
Literature cited ............................................................................................44
Abstract
Phylogenetic relationships of snailfishes of the family Liparidae were analyzed on the basis of two sets of molecular
sequence data: one from the mitochondrial DNA cytochrome c oxidase subunit one gene (COI) and another from
restriction-site associated genome-wide sequences (RADseq). The analysis of COI sequence data from at least 122 species
of 18 genera from the Pacific, Atlantic, and Southern oceans resulted in a moderately well-resolved phylogeny among
the major clades, albeit with significant polytomy among central clades. Nectoliparis was the sister of all other members
of the family, followed by Liparis. Liparis, Careproctus, and Paraliparis were paraphyletic. Liparis was recovered in
two closely related clades, with L. fucensis sister of all other liparids except Nectoliparis, and both Careproctus and
Paraliparis were each recovered among at least three widely separated clades. The RADseq analysis of 26 species of 11
genera from the eastern North Pacific strongly confirmed the overall results of the COI analysis, with the exception of the
paraphyly of Liparis due to the absence of L. fucensis. Our results show that the pelvic disc has been independently lost
multiple times and the pectoral-fin girdle has been independently reduced in multiple lineages.
Key words: Teleostei, molecular, morphology, Liparis, Careproctus, Paraliparis, phylogeny
Introduction
Snailfishes of the family Liparidae are a well-defined group of diverse cottoid fishes characterized most promi-
nently by their scale-less, typically naked, elongate bodies, and by the development of a sucking disk formed by
highly modified pelvic fins. The family is bipolar in distribution and is one of the four most-rapidly speciating
clades of marine fishes, all found in high-latitudes (Rabosky et al. 2018). The Liparidae comprises over 430 species
(Eschmeyer & Fong 2017) allocated to about 32 genera (Chernova et al. 2004; Orr 2004; Balushkin 2012; Stein
2015; Eschmeyer et al. 2017). Two of its most diverse genera, Careproctus and Paraliparis, are recognized in both
the Northern Hemisphere (northern species) and the Southern Hemisphere (southern species), while a third, Liparis,
is restricted to the north. Presently, 214 species are known from the north, primarily the North Pacific, whereas 262
species are recorded from the south, primarily the Southern Ocean, and many new species continue to be described
both north and south (e.g., Chernova 2005ab, 2014; Stein 2006, 2012, 2016; Orr 2102, 2016; Eschmeyer & Fong,
2017). Liparids are particularly diverse in North Pacific waters, especially off Alaska where over 85 species have
either been described or are known but as yet undescribed (Mecklenburg et al. 2002; Orr 2004; Orr & Busby 2006;
Orr & Maslenikov 2007; Orr 2012, 2016; Orr et al. 2015; Orr, unpublished). They are abundant as well as diverse,
as documented in trawl surveys conducted throughout a wide range of depths, especially in surveys of the Aleutian
Islands (Raring et al. 2016) and upper continental slope of the eastern Bering Sea (Hoff 2016).
While bipolar in distribution, and essentially limited to cool-temperate and cold waters, liparids are widely
distributed from the shallowest waters of the intertidal region to near the deepest parts of the ocean. The recently
described Mariana Snailfish, Pseudoliparis swirei, has been documented at depths to 8178 m and is now recognized
as the deepest dwelling vertebrate thus far confirmed (Linley et al. 2016; Gerringer et al. 2017a; JAMSTEC 2017).
In addition, although not yet captured and thus not formally described, the Ethereal Snailfish was filmed in the
Mariana Trench at a depth of 8,145 m, only 33 m shallower (Linley et al. 2016; Gerringer et al. 2017a). Most spe-
cies are demersal or benthic, but many species are found also in the midwater. Their precise contributions to marine
ecosystems are unclear, but liparids are important as prey and as predators in many ecosystems (Bansode et al. 2014;
MOLECULAR PHYLOGENETICS OF THE LIPARIDAE Zootaxa 4642 (1) © 2019 Magnolia Press · 3
Gerringer et al. 2017b; Whitehouse et al. 2017). Among their unusual behavioral characteristics, species of Care-
proctus are known to lay their eggs within the carapace of lithodid king crabs (Stein 1980; Gardner et al. 2016).
Phylogenetic status. Historically placed within the Scorpaeniformes among the sculpins (Nelson et al. 2016),
the Liparidae was recently reallocated to the perciform suborder Cottoidei by Smith & Busby (2014; = infraorder
Cottales of Betancur-R et al. 2017). Its close relationship with the Cyclopteridae, as originally proposed by Peter
Artedi (1738; see also Walbaum, 1792), followed later by Cuvier (1817) and Garman (1892), has been unchal-
lenged, and both families are unquestionably monophyletic (Near et al. 2012; Smith & Busby 2014). While sys-
tematic progress has been made with the Cyclopteridae (Kai et al. 2015; Oku et al. 2017; Stevenson et al. 2017),
the phylogenetic relationships of the far more diverse Liparidae are poorly understood (Smith & Busby 2014) and
the monophyly of major genera within the family has been questioned (Andriashev 1986; Knudsen & Møller 2007;
Balushkin 2012). Previous studies have been limited primarily by inadequate taxon sampling of this morphologi-
cally diverse and species-rich group, as well as by the challenges of working with the morphology of these poorly
ossified, reductive, and easily damaged fishes.
Early works focused on the few species known at the time and gave only general ideas of their evolution and
relationships: Garman (1892) provided the first detailed anatomical descriptions and reviewed all 18 known spe-
cies in the three genera—Liparis, Careproctus, and Paraliparis—recognized at the time; Goode & Bean (1896)
reviewed 10 deep-sea species in five genera; and Jordan & Evermann (1898) included all 39 species known among
10 genera in North American waters. Burke’s (1930) monumental revisionary work on the family summarized all
that was known at the time about the family, its 114 species in 13 genera, and its evolution. As in the works of pre-
vious authors, Burke described the family as being closely related to the Cyclopteridae and more distantly to the
broadly defined Cottidae, supporting the early consensus about evolution within the group: that is, their evolution
from shallow-water, large-disc species of Liparis, through disc reduction among deeper dwelling Careproctus, to
the complete loss of the pelvic disc among deeper dwelling and semi-pelagic species of Paraliparis and related
genera. Thus, genera were arranged in phylogenetic order from least to most derived, with those more basal bearing
a disc (i.e., Liparis and Polypera, Careproctus and Temnocora, Crystallichthys and Crystallias, and Gyrinichthys)
and those more derived lacking or having a highly reduced disc (i.e., Elassodiscus, Paraliparis, Rhinoliparis, Acan-
tholiparis, Nectoliparis, and Lipariscus).
Overall, later analyses supported these general patterns, but some notable exceptions were demonstrated. Es-
pecially as the full diversity of the Liparidae has become more clearly understood, wide gaps in our understanding
of the phylogenetics have been revealed. Several modern phylogenetic analyses have been published based on
morphological and molecular data (Figs. 1–7). As in other complex groups, analyses have moved from those using
strictly morphological data, to combined analyses including both molecular and morphological data, and most re-
cently to strictly molecular data. Taxon and individual sampling have generally increased among species and genera
with each analysis.
The earliest analyses used morphological characters exclusively. In the first modern phylogenetic analysis,
Kido (1988; Fig. 1) examined 60 species in about nine genera using osteology and external morphology, focusing
on species of Japanese waters augmented by selected species from the North Atlantic. In challenging aspects of
Kido’s (1988) results, particularly the position of Nectoliparis as sister of all other liparids, Balushkin (1996; Fig. 2)
attempted to describe the relationship of his new monotypic genus, Palmoliparis, among the other 26 known liparid
genera. While preliminary and supported by only seven osteological and external characters, his analysis was the
first attempt to place all known genera within a cladistic scheme based on well-recognized character transforma-
tions and current interpretations of liparid evolution (Andriashev 1998; Andriashev & Stein 1998). In both of these
morphological analyses, Liparis was monophyletic and Careproctus was paraphyletic (in the reanalysis of Kido’s
1988 data by Knudsen et al. 2007). Paraliparis was also paraphyletic in Kido’s (1988) analysis; its status was not
examined by Balushkin (1996).
The first phylogenetic analysis to use molecular data was that of Knudsen et al. (2007; Fig. 3), who conducted a
combined analysis of two mitochondrial DNA (mtDNA) datasets of over 1000 bp with morphological data, based on
24 species in nine North Atlantic and North Pacific genera. With this limited taxon sampling, they found all genera
except Paraliparis to be monophyletic.
More recent species-level analyses have used only mitochondrial sequence data, primarily the cytochrome c
oxidase subunit I gene (COI), and have focused on the identification of species. As collections of tissues have in-
creased, numbers of species, genera, and individuals have grown with each analysis.
Steinke et al. (2009; Fig. 4) analyzed sequence divergences in a 650 bp alignment of COI of 78 tissues of 19
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species of eight Canadian Pacific genera. Their goal was to evaluate the utility of COI for distinguishing species,
although misidentifications hampered the accuracy of their species-level conclusions. In their neighbor-joining
tree, Liparis was monophyletic and both Paraliparis and Careproctus were paraphyletic. Similarly, Duhamel et al.
(2010; Fig. 5) focused on Antarctic species and significantly increased the overall number of species and geographic
scope sampled by analyzing 157 tissues of 46 species in nine genera. With much new material of Antarctic species
and 103 COI sequences 668 bp in length from other geographic areas taken from online databases, their Bayesian
analysis found not only Paraliparis and Careproctus to be paraphyletic but, for the first time, Liparis as well.
FIGURE 1. Phylogenetic hypothesis of Kido (1988, after fig. 20), derived from a maximum parsimony analysis of morphologi-
cal data, using 34 osteological and external characters, for 60 liparid species.
MOLECULAR PHYLOGENETICS OF THE LIPARIDAE Zootaxa 4642 (1) © 2019 Magnolia Press · 5
FIGURE 2. Phylogenetic hypothesis of Balushkin (1996, after fig. 4), derived from a manual cladistic analysis of morphologi-
cal data, including seven osteological and external characters, for 26 liparid genera.
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FIGURE 3. Majority-rule (50%) consensus tree of Knudsen et al. (2007, after fig. 3), derived from a Bayesian analysis of three
combined datasets composed of mitochondrial DNA (16S and cytochrome b) and morphological data for 24 liparid species. Tree
is rooted with species of the Cyclopteridae. Posterior probabilities are above branches.
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FIGURE 4. Unrooted neighbor-joining tree of Steinke et al. (2009, after fig. 4), derived from cluster analysis of a 650 bp align-
ment of cytochrome c oxidase subunit 1 gene (COI) sequences for 78 samples of 19 liparid species. Bootstrap values >80 are
above branches leading to multiple species. Corrected identifications based on our study are in parentheses.
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FIGURE 5. Consensus phylogenetic tree of Duhamel et al. (2010, after fig. 3), derived from Bayesian and maximum parsimony
analyses of a 668 bp alignment of cytochrome c oxidase subunit 1 gene (COI) sequences for 157 samples of 46 liparid species.
Bayesian posterior probabilities are above branches that lead to multiple species. Tree is rooted with species of the Cyclopteri-
dae and Zoarcidae. Corrected identifications based on our study are in parentheses.
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FIGURE 6. Majority rule (50%) consensus phylogenetic tree of Gardner et al. (2016, after fig. 4), derived from Bayesian infer-
ence and maximum parsimony analysis of a 492 bp alignment of cytochrome c oxidase subunit 1 gene (COI) sequences of 492
bp for 128 samples of 23 liparid species. Bootstrap values are above and Bayesian posterior probabilities are below branches that
lead to multiple species. Tree is rooted with Liparis gibbus. Corrected identifications based on our study are in parentheses.
Primarily concerned with identifying eggs of snailfishes found within lithodid crabs, Gardner et al. (2016; Fig.
6) conducted a Bayesian analysis of 429 bp of COI data for 23 species and nine genera of 53 liparid adults from the
Bering Sea, Aleutian Islands, and eastern North Pacific Ocean. In their analysis, multiple clades of Careproctus and
Paraliparis, as well as other genera, were placed in an unresolved polytomy of their consensus tree.
Most recently, Shen et al. (2017; Fig. 7) published the full mitochondrial genome of an undescribed deep-water
liparid and attempted to place the species phylogenetically through the analysis of mtDNA data, including COI,
taken from online databases. In analyses with the 16S and Cytb mtDNA data, their overall results were unsurpris-
ingly similar to those of Knudsen et al. (2007), the primary source of their data. With the COI data alone for 440 bp
of 83 species in nine genera, they produced a tree that resolved all three major genera as paraphyletic.
Methodological approach. Despite these many previous efforts, the phylogenetic relationships of liparids
remain poorly understood and increasing evidence reveals incongruence between liparid taxonomy and its phylo-
genetic history. The explosion of readily available molecular data, especially the wide use of COI sequence data
stored in public databases, and the development of new next-generation DNA sequencing techniques, which incor-
porate more data than typically employed by Sanger sequencing techniques and encompass a greater range of sites
throughout the genome, have provided new resources and tools for phylogenetic research (e.g., Leaché et al. 2015).
The time is thus ripe for more extensive molecular phylogenetic analyses.
The mitochondrial COI gene is relatively easy to sample and because of widespread efforts to routinely sample
species to capture biodiversity information, copious data are available from public databases, such as the Barcode of
Life Database (BOLD; Ratnasingham & Hebert 2007) and GenBank (Benson et al. 2005). However, the gene may
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